Transcription of DNA into messenger RNA is the fundamental step in cellular metabolism that allows an organism to express its genetic information. Transcription is regulated by the needs of the organism. This control is in many cases exerted by factors or molecules that influence the ability of the transcribing enzymes, RNA polymerase, to initiate RNA synthesis; in so far as is known, such factors influence the rate of stability of binding of RNA polymerase to promoter sites. The research proposed in this application has its objective characterization of some aspects of the binding reaction and the subsquent initiation reaction. Most previous work in this area has focussed on the properties of Escherichia coli RNA polymerase. We propose to extend detailed analysis of promoter-polymerase interaction to RNA polymerases from bacillus subtilis and Streptomyces coelicolor, two gram-positive, differentiating bacteria. Our experimental approach will be to look for contacts between RNA polymerase and specific DNA bases of well defined promoter sites. In corollary experiments we will examine the function of the Sigma subunit of RNA polymerase in the early steps of RNA synthesis. By the use of plasmids we have constructed in which the lac genes of E. coli have been brought under the control of a B. subtilis promoter, we will isolate and characterize mutants with promoter activity.